Acute, infectious diarrhoea is a leading cause of disease and death in many areas of the world. In developing countries, the impact of diarrhoeal disease is very important. For Asia, Africa and Latin America, it has been estimated that there are between 3-4 billion cases of diarrhoea each year and of those cases about 5-10 million result in death (Walsh, J. A. et al.: N. Engl. J. Med., 301:967-974 (1979)).
Rotaviruses have been recognised as one of the most important causes of severe diarrhoea in infants and young children (Estes, M. K. Rotaviruses and Their Replication in Fields Virology, Third Edition, edited by Fields et al., Raven Publishers, Philadelphia, 1996). It is estimated that rotavirus disease is responsible for over 600,000 deaths annually. Rotavirus-induced illness most commonly affects children between 6 and 24 months of age, and the peak prevalence of the disease generally occurs during the cooler months in temperate climates, and year-round in tropical areas. Rotaviruses are typically transmitted from person to person by the faecal-oral route with an incubation period of from about 1 to about 3 days. Unlike infection in the 6-month to 24-month age group, neonates are generally asymptomatic or have only mild disease. In contrast to the severe disease normally encountered in young children, most adults are protected as a result of previous rotavirus infection so most adult infections are mild or asymptomatic (Offit, P. A. et al. Comp. Ther., 8(8):21-26, 1982).
Rotaviruses are spherical, and their name is derived from their distinctive outer and inner or double-shelled capsid structure. Typically, the double-shelled capsid structure of a rotavirus surrounds an inner protein shell or core that contains the genome. The genome of a rotavirus is composed of 11 segments of double-stranded RNA which encode at least 11 distinct viral proteins. Two of these viral proteins designated as VP4 (P protein) and VP7 (G protein) are structural proteins arranged on the exterior of the double-shelled capsid structure. The inner capsid of the rotavirus presents one protein, which is the rotavirus protein designated VP6. The relative importance of these three particular rotavirus proteins in eliciting the immune response that follows rotavirus infection is not yet clear. Nevertheless, the VP6 protein determines the group and subgroup antigen, and VP4 and VP7 proteins are the determinants of serotype specificity.
To date, at least 14 rotavirus G serotypes and 11 rotavirus P serotypes have been identified (Linhares A. C. & Bresse J. S., Pan. Am. J. Publ. Health 2000, 9, 305-330). Among these, 10 G serotypes and 6 P serotypes have been identified among the human rotavirus.
VP7 protein is a 38,000 MW glycoprotein (34,000 MW when non-glycosylated) which is the translational product of genomic segment 7, 8 or 9, depending on the strain. This protein stimulates formation of the major neutralising antibody following rotavirus infection. VP4 protein is a non-glycosylated protein of approximately 88,000 MW which is the translational product of genomic segment 4. This protein also stimulates neutralising antibody following rotavirus infection. Since VP4 and VP7 proteins are the viral proteins against which neutralising antibodies are directed, they are believed to be prime candidates for development of rotavirus vaccines, affording protection against rotavirus illness.
Natural rotavirus infection during early childhood is known to elicit protective immunity.
Early vaccine development for preventing rotavirus infections began in the 1970s after the discovery of the virus. Initially, attenuated strains from animals and humans were studied, whilst more recent efforts have focused on human-animal reassortants.
The development of novel rotavirus formulations must comply with a number of requirements, including worldwide distribution potential and stability under a broad range of environmental and storage conditions. In particular, the stability of a formulation, especially of a pharmaceutical or vaccine composition, will in general be better at lower temperatures compared to room or higher temperatures.
Consequently one stabilisation method has been to develop vaccine formulations that can be stored frozen (−20° C. to −70° C.) or alternatively to develop lyophilised vaccines that can be kept for a prolonged period of time at around refrigerator temperature (2° C. to 8° C.). However, it is a known fact that the lyophilisation process has a limiting capacity, and is associated with a high production cost. Furthermore, lyophilised vaccines have a more sophisticated handling for administration as they may require more complex, hence relatively expensive devices such as multichamber/vial vaccines, with the active ingredient in one chamber and the reconstitution liquid in another chamber. Lyophilised vaccines are also associated with higher shipment and storage cost. These options may be inadequate for some countries in the developing world where the administration device has to be financially affordable and where the availability of production and storage infrastructure may be inexistent or unreliable.
As Rotavirus are conventionally administered orally to human infants, this route brings several challenges to immunogenic rotavirus compositions.
Rotavirus is rapidly inactivated in an acidic environment, upon exposure to acid buffer or acidic gastric juice for example (C. Weiss and H. F. Clark, 1985, J. Gen. Virol., 66, 2725-2730; T. Vesikari et al., 1984, The Lancet, page 700; R. H. Foster and A. J. Wagstaff, 1998, BioDrugs February: 9(2) 155-178). Therefore it is desirable that rotavirus compositions are formulated in a way that they are stable during storage and after administration into the host recipient.
Rotavirus vaccines are primarily intended to be administered to babies, as early as at the age of 4 weeks. A small vaccine dose volume, such as lower than 2 ml or even than 1.5 ml dose volume, will be advantageous for that population. Therefore, it is desirable that rotavirus compositions are formulated in a small dose volume.
Stabilising formulations for liquid viral vaccines are known. For example, EP 0 065 905 discloses in general stabilising compositions suitable for a series of viruses such as those causing measles or influenza, and in particular it discloses stabilizing phosphate buffer—containing solutions suitable for live attenuated virus.
Other stabilizing formulations are disclosed in WO 98/13065 and in Clark et al. (Pediatr Infect Dis J. 2003 October; 22(10):914-20). Such formulations also require, amongst other constituents, the presence of phosphate to act as a buffering agent to neutralise stomach acidity. These formulations are however not compatible with the requirements set out above for the successful development of a rotavirus formulation, specifically they are not compatible with a reduced volume of the vaccine dose that is best suited for a human infant. In particular, the present inventor has found that adapting this prior art formulation into a low volume setting such as 1.5 ml or lower, whilst maintaining efficient antacid capacity, leads to problems arising from inappropriate concentration of the formulation constituents, in particular phosphate buffer.
There is a need therefore to develop alternative rotavirus formulations, in particular alternative liquid formulations that can withstand gastric acidity, and are refrigerator-stable despite the absence of phosphate. In addition there is a need that such alternative formulations be also successfully formulated in a vaccine dose volume as small as possible.
Therefore the present invention not only provides alternative stable immunogenic compositions that are devoid of phosphate or contain only minimal amounts of phosphate, but also allow rotavirus to be formulated in a low dose volume that are suitable for oral administration to human infants.